Profiling of BABA-induced differentially expressed genes of Zea mays using suppression subtractive hybridization

Abstract

β-Aminobutyric acid (BABA), a potent chemical priming agent, induces plant resistance to a broad spectrum of biotic and abiotic stresses. Nevertheless, the molecular mechanisms underlying this phenomenon remain poorly understood. Therefore, this study aims to identify rare and differentially expressed transcripts in BABA-primed maize (Zea mays L.) using a suppression subtractive hybridization strategy. Forward subtraction was performed using the cDNA prepared from a BABA primed leaf as a tester and cDNA from the control leaf as the driver. For reverse subtraction, tester cDNA was synthesized from control leaf and driver cDNA from the BABA primed leaf. A total of 626 clones were picked randomly. In total 192 and 204 high quality expressed sequence tags (ESTs) were generated from forward (ZmSF) and reverse SSH library (ZmSR) respectively. Contigs analyses revealed 27 unigenes from the forward subtracted library and 4 unigenes from the reverse subtracted library. The ESTs encoding alpha amylase, inositol monophosphatase 3, tocopherol cyclase, thioredoxin H-type, aspartic proteinase, NADP-malic enzyme and PEP carboxykinase were expressed predominantly in BABA-primed leaf and found to be predominantly associated with major metabolic and biosynthesis events along with seed germination, developmental processes and adaptive responses to environmental stimuli such as drought, salinity, osmotic stress and pathogen attack. In this study, we present the first report of transcriptome analysis of maize leaf exposed to BABA. The expression profiles of candidate genes from subtracted libraries were checked using semi quantitative RT-PCR and further validated by qRT-PCR. Expression profiles largely corroborated the quality and functional diversity of both libraries. In ZmSF, 130 ESTs (67.70%) did not show any significant similarity to any protein in the NCBI database, indicating a significant fraction of novel and rare expressed transcripts that were rapidly and strongly induced in response to BABA. Further research needs to be done for these novel transcripts to elucidate their function in a BABA-potentiated stress adaptation mechanism. Overall, our study adds new insights into the BABA potentiated defense mechanisms in plants, and strengthens the available transcriptome database of maize.